The development and maintenance of the vertebral skeleton is a complex process, but in simplest terms represents a balance between osteoblast-induced mineralization and osteoclast-induced demineralization. Osteoblast-like cells are also present in the vascular wall and participate in the earliest manifestations of atherosclerosis. Hydroxyapatite (HA; Ca10(PO4)6(OH)2, also know as hydroxylapatite) is the major mineral product of osteoblasts and calcifying vascular cells and binds naturally occurring pyrophosphates and phosphonates with high affinity. Osteoblastic activity occurs at sites of new bone formation, i.e., sites of deposition of hydroxyapatite. New bone formation occurs after fracture, at sites of bony infections, and especially at the sites of certain cancer metastases (e.g., prostate cancer metastases).
At the present time, osteoblastic activity is detected in vivo using radionuclides and SPECT imaging. For instance, a common technique is the “bone scan”, which utilizes the radiometal 99mTc coupled to the bisphosphonate compound methylene bisphosphonate (MDP). Unfortunately, radioscintigraphic detection does not provide high-resolution anatomical detail, and requires the use of radioactive compounds.
Fluorescence imaging is found at the heart of numerous chemical and biomedical analysis schemes. Many of these schemes are based on the introduction of a fluorescent species as a marker, stain, dye or indicator (Devoisselle et al. (1993) Optical Engineering 32:239; Haugland and Minta, “Design and Application of Indicator Dyes,” Noninvasive Techniques in Cell Biol., ed. B. H. Satir, Chap: 1, p 1, (Wiley-Liss, New York, N.Y., 1990); Gross, “Quantitative Single Cell Fluorescence Imaging of Indicator Dyes,” Noninvasive Techniques in Cell Biol., ed. B. H. Satir, Chap. 2, p 21, (Wiley-Liss, New York, N.Y., 1990). To date, however there has not been a non-isotopic method for directly detecting HA in vivo.
It is an object of the present invention to use bisphosphonate compounds for use in non-isotopic (i.e., without the need for radioactivity) detection of hydroxyapatite, such as to determine osteoblastic activity in vivo.